USP18 in Cancer Development The long-term goal of this study is to understand the role of USP18 in cancer development and targeted therapy. USP18 is a member of the ubiquitin specific protease family. We initially cloned genes encoding mouse and human USP18 as UBP43 (ubiquitin protease with 43 kDa) during a study of a leukemia fusion protein. Our further analyses revealed that the major protease activity of USP18 is removal of a ubiquitin like modifier ISG15 from ISGylated proteins and that USP18 is a potent inhibitor of Type I interferon (IFN) signaling independent of its ISG15 deconjugating enzyme activity. Moreover, studies from other groups and our own lab demonstrate that USP18 regulates cancer development. We report that USP18 deficiency impairs development of BCR-ABL induced chronic myeloid leukemia in a retrovirus infection mediated hematopoietic stem cell transplantation mouse model and slows down mammary tumor growth in a papilloma middle tumor- antigen (PyVmT) transgenic mouse model. Interestingly, we also discovered that in addition to blocking Type I IFN signaling, USP18 inhibits Type III IFN effects. This finding is highly significant since the Type III IFN receptor is mainly expressed in solid tumor initiating epithelial cells but not in IFN inducible inflammatory cytokine producing blood cells. However, the molecular mechanism of USP18 in Type III IFN signaling is unclear. Our recent unpublished data demonstrate that 1) knockout of the Usp18 gene specifically in myeloid cells slows down tumor growth in three tested mouse cancer models (B16 melanoma, EL4 thymoma, and LLC lung carcinoma), 2) lack of Usp18 expression in MLL-AF9 and RUNX1-ETO9a fusion protein induced acute myeloid leukemia initiation cells slows down leukemia development and activates both the DNA damage mediated cell response pathway and the Type I IFN signaling pathway. Therefore, this proposal will test the hypothesis that USP18 and its downstream effectors are potential therapeutic targets due to its role in regulating signaling pathways of cancer cells and non-cancer myeloid cells in the cancer microenvironment. We propose to address molecular mechanisms of USP18 in cancer development with the following specific aims: 1) Understanding the molecular basis of USP18 in Type III IFN signaling, 2) Examine the role of USP18 in modulating tumor associated myeloid cells, 3) Analyze the role of USP18 in leukemia initiating cells. The proposed studies are based on our accumulated knowledge and our most recent novel findings on USP18. This proposal will address important questions about molecular mechanisms of USP18 in cancer development and may provide novel insights into the prevention and therapeutic treatment of human cancer.